1.1 Field of the Invention
This invention relates to the field of stem cell culture media and to methods for culturing such cells. More particularly, the present invention provides methods and materials for culturing primate-derived primordial stem cells in a substantially undifferentiated state with and without a feeder layer. The present invention has applications in the areas of cell culture, tissue transplantation, drug discovery, and gene therapy.
1.2 The Related Art
Stem cells are cells capable of differentiation into other cell types having a particular, specialized function (“fully differentiated” cells) or other types of stem cells which are capable of differentiation into a more narrow range of cell types (“pluripotent” cells). Stem cells having the ability to differentiate into any type of cell, i.e., pluripotent or fully differentiated, are called “totipotent”. Such cells are also referred to as “primordial stem cells”. There has been great interest in isolating and growing primordial stem cells from primates, especially from humans, as such primordial stem cells could provide a supply of readily available cells and tissues of all types for use in transplantation, drug discovery, and gene therapy in humans. Methods for isolating and growing primordial stem cells from primates have been described. Procedures for isolating and growing human primordial stem cells are described in co-pending U.S. patent application Ser. No. 08/829,372. Procedures for obtaining Rhesus monkey and other non-human primate primordial stem cells are described in co-pending U.S. patent applications Ser. Nos. 08/376,327; 08/591,246; 08/665,217; and WO 96/22362. Each of these patent applications is incorporated herein by reference in its entirety and for all purposes. In addition, methods for isolating Rhesus monkey primordial stem cells are described in Thomson et al. (1995 Proc. Natl. Acad. Sci. USA 92:7844-7848) also incorporated herein by reference in its entirety and for all purposes.
Unfortunately, current methods for growing primordial stem cells derived from primates in culture have not been as clearly defined as, and are relatively inefficient compared with, methods for culturing primordial stem cells for other species such as mouse. For example, current methods of culturing primate-derived primordial stem cells require a feeder layer that complicates and slows the process of cell cultivation. In addition, the formulation of an optimal culture media for propagating undifferentiated totipotent primate-derived primordial stem cells remains to be determined.
In particular, it is desirable to maintain cultures of totipotent primordial stem cells for extended periods or indefinitely. The ability to maintain cultures of undifferentiated, totipotent, primate-derived primordial stem cells for long periods facilitates the use of such cells for therapeutic purposes. Moreover, it would be desirable to grow cultures of substantially undifferentiated primate-derived primordial stem cells for periods sufficient to allow the production of primate-derived primordial stem cells having multiple genetic modifications for the production of tissues and for gene therapy.